Water-wettable chromatographic media for solid phase extraction

ABSTRACT

A method for removing an organic solute from a solution, comprising contacting the solution with a polymer formed by copolymerizing one or more hydrophobic monomers and one or more hydrophilic monomers, whereby the solute is adsorbed onto the polymer. The solution can comprise a polar solvent such as a polar organic solvent or water or an aqueous buffer. The hydrophobic monomer can be, for example, divinylbenzene. The hydrophilic monomer can be, for example, a heterocyclic monomer, such as a vinylpyridine or N-vinylpyrrolidone.

RELATED APPLICATIONS

This application is a continuation application of U.S. Ser. No.09/216,047, filed on Dec. 18, 1998, now U.S. Pat. No. 5,976,367, whichis a divisional application of U.S. Ser. No. 08/634,710 filed on Apr.18, 1996, now U.S. Pat. No. 5,882,521. The teachings of each of thesereferenced applications arc expressly incorporated herein by referencein their entirety.

BACKGROUND OF THE INVENTION

Solid phase extraction is a chromatographic technique of frequent use inthe preparation of samples for quantitative analysis, for example, viahigh performance liquid chromatography (HPLC) or gas chromatography (GC)(McDonald and Bouvier, eds. Solid Phase Extraction Applications Guideand Bibliography, sixth edition, Milford, Mass.: Waters (1995)). Solidphase extraction can be used to separate a component of interest in acomplex solution from potentially interfering matrix elements and toconcentrate the analyte to levels amenable to detection and measurement.Thus, solid phase extraction is of use in the analysis of environmentalsamples, where, for example, various soluble components of soils mayinterfere with the analysis of trace organic materials. Solid phaseextraction is also of importance in the analysis of pharmaceuticalagents or metabolites in blood plasma, which requires the prior removalof plasma proteins and other matrix constituents which may interferewith the analysis.

Solid phase extraction of an aqueous solution is typically performed bypassing the solution through a single-use cartridge containing achromatographic sorbent. The most commonly used sorbents consist ofporous silica particles that have been functionalized on their surfacewith hydrophobic octyl (C₈) and octadecyl (C₁₈) functional groups. Priorto use, such sorbents must be wetted with a water-miscible polar organicsolvent to solvate the alkyl chains. This increases the contact of thesechains with the aqueous phase, increasing the sorbent surface areaavailable to solutes and, therefore, retention of solutes. Such sorbentswhich are not pre-wetted or have dried out display poor soluteretention, and, thus, inadequate separation of solution components.

The requirement that the sorbent remain wetted during the extractionprocedure complicates solid phase extractions and substantially slowssample analysis. For example, solid phase extraction cartridges, ingeneral, have differing flow rates and must be monitored individually toprevent drying out when used on a vacuum manifold, the current state ofthe art for processing multiple samples. This further complicates thedevelopment of instruments for automated solid phase extraction, whichoften incorporate elaborate safeguards to prevent drying out of thesorbent.

Thus, there is need for a solid phase extraction method which utilizes asorbent that does not require wetting with an organic solvent or thatstays wetted even if the bulk of the wetting solvent is removed duringuse on a vacuum manifold. Such a method would enable more rapid samplepreparation for quantitative analysis, particularly for multiplesamples, and allow the development of less expensive and simpler methodsfor automated solid phase extraction.

SUMMARY OF THE INVENTION

The present invention relates to a method for removing an organic solutefrom a solution. The method comprises contacting the solution with awater-wettable polymer formed by copolymerizing one or more hydrophobicmonomers and one or more hydrophilic monomers, whereby the solute isadsorbed onto the polymer. The solution can comprise a polar solventsuch as a polar organic solvent, a water/organic mixture or, preferably,water or an aqueous solution, such as an aqueous buffer, acid, base orsalt solution.

The hydrophobic monomer can comprise a hydrophobic moiety. Suitablehydrophobic moieties include, but are not limited to phenyl, phenyleneand C₂ -C₁₈ -alkyl groups. Suitable hydrophobic monomers includedivinylbenzene and styrene.

The hydrophilic monomer can comprise a hydrophilic moiety. In oneembodiment the hydrophilic moiety is a saturated, unsaturated oraromatic heterocyclic group, such as a pyrrolidonyl group or a pyridylgroup. In another embodiment, the hydrophilic moiety is an ether group.Suitable hydrophilic monomers are, for example, N-vinylpyrrolidone,2-vinylpyridine, 3-vinylpyridine, 4-vinylpyridine and ethylene oxide.

In one embodiment of the method, the polymer is apoly(divinylbenzene-co-N-vinylpyrrolidone) copolymer which comprisesgreater than about 12 mole percent N-vinylpyrrolidone. In a preferredembodiment, the copolymer comprises from about 15 mole percent to about30 mole percent N-vinylpyrrolidone.

The present invention further includes a method for forming a solution,containing a solute, which is suitable for quantitative analysis. In oneembodiment, the method comprises contacting a first solution includingthe solute with a water-wettable polymer formed by copolymerizing atleast one hydrophobic monomer and at least one hydrophilic monomer,whereby the solute is adsorbed onto the polymer. This is followed bywashing the polymer with a suitable solvent or mixture of solvents, sothat the solute is desorbed from the polymer, thereby forming a secondsolution including the solute. This second solution is suitable forquantitative analysis.

In another embodiment, the invention provides a method for forming asolution comprising a polar organic solute which is suitable forquantitative analysis. The method comprises contacting a solution whichincludes the polar organic solute and at least one additional solute oflesser polarity with a water-wettable polymer formed by copolymerizingat least one hydrophobic monomer and at least one hydrophilic monomer,whereby the additional solute is adsorbed onto the polymer and the polarsolute remains in the aqueous phase. The resulting aqueous phase is,thus, a solution of the polar organic solute which is suitable forquantitative analysis.

The present invention further includes a solid phase extractioncartridge comprising an open-ended container and a polymer packed withinthe container. The solid phase extraction cartridge can, optionally,further comprise a porous retaining means, such as a frit. The polymeris formed by copolymerizing at least one hydrophobic monomer and atleast one hydrophilic monomer. Suitable polymers includepoly(divinylbenzene-co-N-vinylpyrrolidone) copolymers which compriseabout 12 mole percent or more, preferably from about 15 mole percent toabout 30 mole percent, N-vinylpyrrolidone. The solid phase extractioncartridge preferably comprises from about 0.025 g to about 1 g of thepolymer.

The present invention enables the solid phase extraction of one or moresolutes from an aqueous solution, without prior wetting of the sorbentwith an organic solvent. The method is versatile with respect to soluteidentity, resulting in extraction of a broad range of solutes of varyingpolarity. A particular advantage of the method is that the sorbent candry out during the extraction procedure without diminishing the abilityof the sorbent polymer to retain solutes. Thus, the present inventionprovides a simpler method for the preparation of analytical samples,decreasing sample preparation time and increasing sample throughput. Thepresent method is, thus, also more amenable to automation than currentlyused methods.

BRIEF DESCRIPTION OF THE DRAWING

The FIGURE is a schematic, in cross-section, of one embodiment of thesolid phase extraction cartridge of the present invention.

DETAILED DESCRIPTION OF THE INVENTION

The present invention provides a method for solid phase extraction ofaqueous or buffered aqueous solutions which does not require that thesorbent be wetted with an excess of organic solvent prior to and duringthe solid phase extraction process. The invention is based on thediscovery that polymers or resins comprising both a hydrophilic monomerand a hydrophobic monomer in a suitable ratio can be wetted by waterwhile maintaining surprisingly effective retention of organic soluteswith a wide range of chromatographic polarities.

As described in the Exemplification, a relatively small increase in theN-vinylpyrrolidone content of apoly(divinylbenzene-co-N-vinylpyrrolidone) copolymer resulted in adramatic improvement in retention of polar organic solutes underconditions in which the pre-wetted polymer was dried under rcducedpressure for several minutes. For example, under these conditions,recovery of acetaminophen from such a copolymer comprising 9 molepercent N-vinylpyrrolidone was 10.4%. Increasing the mole percentN-vinylpyrrolidone in the copolymer to 13 resulted in a 92% recovery ofacctaminophen. Similar results were observed for procainamide,ranitidine, and caffeine. For relatively nonpolar solutes the differencein recovery between the two copolymers was less dramatic.

The ability of poly(divinylbenzene-co-N-vinylpyrrolidone) copolymerscomprising between 13 mole percent and 22 mole percentN-vinylpyrrolidone to retain organic solutes was also compared with thatof octadecyl (C₁₈)-bonded silica gel. As discussed in theExemplification, the C₁₈ -bonded silica sorbent showed poor retention ofpolar organic solutes when the sorbent was pre-wetted with an organicsolvent and then dried under reduced pressure prior to extraction. Forexample, this sorbent showed a 2.8% recovery of m-toluamide under theseconditions. In contrast, the poly(divinylbenzene-co-N-vinylpyrrolidone)copolymer comprising 13 mole percent N-vinylpyrrolidone displayed a96.3% recovery of m-toluamide under similar conditions. Overall, theresults demonstrate that a balance between the mutually exclusiveproperties of water-wettability and retention of organic solutes can beachieved in a copolymer which has a suitable ratio of hydrophilicmonomers and hydrophobic monomers.

In one embodiment, the invention is a method for removing a solute froma solution. The method comprises the step of contacting the solutionwith a water-wettable polymer formed by copolymerizing at least onehydrophobic monomer and at least one hydrophilic monomer, whereby thesolute is adsorbed onto the polymer. The solution can comprise water, ora mixture of water and a water-miscible polar organic solvent such asmethanol, ethanol, N,N-dimethylformamide, dimethylsulfoxide oracetonitrile. The solution can also comprise a mixture of water or anaqueous buffer and a polar, water-miscible organic solvent. In aparticularly preferred embodiment, the solution is an acidic, basic orneutral aqueous or predominately aqueous, i.e., greater than about 50%water by volume, solution. The solute is preferably an organic compound.

The solution can be contacted with the polymer in any fashion whichpermits intimate contact of the polymer and the solution, such as abatch or chromatographic process. For example, the solution can beforced through a porous polymer column, disk or plug, or the solutioncan be stirred with the polymer, such as in a batch-stirred reactor. Thesolution can also be added to a polymer-containing well of a microtiterplate. The polymer can take the form of, for example, beads or pellets.The solution is contacted with the polymer for a time period sufficientfor the solute of interest to substantially adsorb onto the polymer.This is typically the time necessary for the solute to equilibratebetween the polymer surface and the solution. The adsorption orpartition of the solute onto the polymer can be partial or complete.

A preferred polymer for use in the present method is water-wettable andhas the ability to retain a variety of solutes of varying polarity. Theterm "water-wettable", as used herein, describes a material which issolvated, partially or completely, by water. The material, thus, engagesin energetically favorable or attractive interactions with watermolecules. These interactions increase the amount of surface area of thematerial which, upon contact with water, is accessible to watermolecules, and, hence, to solutes present in aqueous solution.

The term "monomer", as used herein, refers to both a molecule comprisingone or more polymerizable functional groups prior to polymerization, anda repeating unit of a polymer. A polymer can comprise two or moredifferent monomers, in which case it can also be referred to as acopolymer. The "mole percent" of a given monomer which a copolymercomprises is the mole fraction, expressed as a percent, of the monomerof interest relative to the total moles of the various (two or more)monomers which compose the copolymer.

In one embodiment of the method, the solution is contacted with thepolymer in dry form. In another embodiment the polymer is wetted priorto contacting the solution with the polymer, for example, by treatingthe polymer with a polar organic solvent, followed by water or anaqueous buffer.

The hydrophilic monomer can comprise hydrophilic group. In oneembodiment, the hydrophilic group is a heterocyclic group, for example,a saturated, unsaturated or aromatic heterocyclic group. Suitableexamples include nitrogen-containing heterocyclic groups such aspyrrolidonyl and pyridyl groups. In another embodiment, the hydrophilicmoiety is an ether group. The hydrophilic monomer can be, for example,N-vinylpyrrolidone, 2-vinylpyridine, 3-vinylpyridine, a hydrophobicmoiety, 4-vinylpyridine or ethylene oxide.

The hydrophobic monomer can comprise, for example, an aromaticcarbocyclic group, such as a phenyl or phenylene group, or an alkylgroup, such as a straight chain or branched C₂ -C₁₈ -alkyl group.Suitable hydrophobic monomers include, but are not limited to, styreneand divinylbenzene.

In a preferred embodiment, the polymer to be contacted with the solutionis a poly(divinylbenzene-co-N-vinylpyrrolidone) copolymer. The polymercan comprise about 12 mole percent or more N-vinylpyrrolidone. In aparticularly preferred embodiment, the polymer comprises from about 15mole percent to about 30 mole percent N-vinylpyrrolidone.

The polymer can be in the form of, for example, beads having a diameterin the range from about 5 to about 500 pm, preferably from about 20 toabout 200 μm. The copolymer, preferably, has a specific surface area inthe range from about 200 to about 800 square meters per gram and poreshaving a diameter ranging from about 0.5 nm to about 100 nm.

The solution comprising the solute can, optionally, further contain oneor more additional solutes. In one embodiment, the solution is anaqueous solution which includes a complex variety of solutes. Solutionsof this type include blood plasma, urine, cerebrospinal fluid, synovialfluid and other biological fluids, including extracts of tissues, suchas liver tissue, muscle tissue, brain tissue and heart tissue. Suchextracts can be aqueous extracts or organic extracts which have beendried and subsequently reconstituted in water or in a water/organicmixture.

The solution can also be ground water, surface water, drinking water oran aqueous or organic extract of an environmental sample, such as a soilsample. The solution can further be a food substance, such as a fruit orvegetable juice or milk or an aqueous or aqueous/organic extract of afood substance, such as a fruit, vegetable, cereal, or meat.

The solute can be any organic compound of polarity suitable foradsorption onto the polymer. Such solutes can include, for example,drugs, pesticides, herbicides, toxins and environmental pollutantsresulting from the combustion of fossil fuels or other industrialactivity, such as metal-organic compounds comprising a heavy metal suchas mercury, lead or cadmium. The solutes can also be metabolites ordegradation products of the foregoing materials. The solutes can alsoinclude biomolecules, such as proteins, peptides, hormones,polynucleotides, vitamins, cofactors, metabolites, lipids andcarbohydrates.

In one embodiment of the method, the polymer is packed as particleswithin an open-ended container to form a solid phase extractioncartridge. The container can be, for example, a cylindrical container orcolumn which is open at both ends so that the solution can enter thecontainer through one end, contact the polymer within the container, andexit the container through the other end. The polymer can be packedwithin the container as small particles, such as beads having a diameterbetween about 5 μm and about 500 μm, preferably between about 20 μm andabout 200 μm. The polymer particles can also be packed in the containerenmeshed in a porous membrane.

The container can be formed of any material which is compatible, withinthe time frame of the extraction process, with the solutions andsolvents to be used in the procedure. Such materials include glass andvarious plastics, such as high density polyethylene and polypropylene.In one embodiment, the container is cylindrical through most of itslength and has a narrow tip at one end. One example of such a containeris a syringe barrel.

The solid phase extraction cartridge can further comprise a porousretaining means, such as a filter element, or frit, at one or both endsof the cartridge adjacent to the polymer to retain the polymer withinthe cartridge and to remove undissolved solid materials from thesolution as it flows into the cartridge, while still permitting solutionflow into and out of the cartridge. Such a filter can be formed from,for example, fritted glass or a porous polymer, such as a porous highdensity polyethylene.

The amount of polymer within the container is limited by the containervolume and can range from about 0.001 g to about 50 g, but is preferablybetween about 0.025 g and about 1 g. The amount of polymer suitable fora given extraction depends upon the amount of solute to be adsorbed, theavailable surface area of the polymer and the strength of theinteraction between the solute and the polymer. This can be readilydetermined by one of ordinary skill in the art.

The present invention includes a solid phase extraction cartridge asdescribed above, wherein the polymer is a water-wettable polymer formedby copolymerizing at least one hydrophobic monomer and at least onehydrophilic monomer. The polymer can be, for example, apoly(divinylbenzcne-co-N-vinylpyrrolidone) copolymer comprising about 12mole percent or more N-vinylpyrrolidone. In a preferred embodiment, thecopolymer comprises from about 15 mole percent to about 30 mole percentN-vinylpyrrolidone. The cartridge can be a single use cartridge, whichis used for the treatment of a single sample and then discarded, or itcan be used to treat multiple samples.

A preferred embodiment of the solid phase extraction cartridge of thepresent invention is illustrated in cross section in the FIGURE.Container 1 is a syringe barrel which can be formed of moldedpolypropylene and can have a volume ranging from about 1 cm³ to about 50cm³. Water wettable polymer 2 is prepared by the copolymerization ofN-vinylpyrrolidone and divinylbenzene and comprises from about 12 molepercent to about 30 mole percent N-vinylpyrrolidone. Polymer 2 is packedwithin the container as porous beads of diameter between about 20 gm andabout 200 μm. The mass of polymer 2 packed within the container canrange from about 0.025 g to about 10 g, depending upon the volume of thecontainer. Frits 3 and 4 are formed of porous high density polyethylene.

The solution to be treated is added to the top of the solid phaseextraction cartridge and allowed to flow through the cartridge, bringingthe solute to be adsorbed into contact with the polymer. The solutioncan flow through the cartridge under the force of gravity. Increasedflow rates can be achieved by establishing a pressure difference betweenthe ends of the cartridge. Such a pressure difference can be establishedby attaching a vacuum source to the lower end of the cartridge or byapplying positive pressure to the upper end of the cartridge, forexample, by applying a pressurized gas, such as air or nitrogen, to thetop of the cartridge, or by compressing the air within the cartridgeabove the polymer with a piston or plunger. The flow rate of thesolution through the cartridge can be adjusted by regulating thepressure difference across the cartridge. Suitable solution flow rates,given in terms of the linear velocity of the solution, range up to about14 mm/second, but are preferably in the range from about 0.7 to about3.5 mm/second.

Another aspect of the present invention is a method for forming asolution of a solute which is suitable for quantitative analysis. In oneembodiment, the solute is of a polarity suitable for adsorption onto thepolymer. The method comprises contacting a first solution which includesthe solute with a polymer formed by copolymerizing at least onehydrophobic monomer and at least one hydrophilic monomer, whereby thesolute is adsorbed onto the polymer. This is followed by washing thepolymer with a suitable, stronger solvent or mixture of solvents,thereby desorbing or eluting the solute from the polymer and forming asecond solution which contains the solute. This second solution issuitable for the quantitative analysis of the solute.

The solution contacted with the polymer can comprise the solute ofinterest in dilute form, for example, at a concentration too low foraccurate quantitation. By adsorbing the solute onto the polymer and thendesorbing the solute with a substantially smaller volume of a less polarsolvent, a solution which includes the solute of interest can beprepared having a substantially higher concentration of the solute ofinterest than that of the original solution. The method also results insolvent exchange, that is, the solute is removed from a first solventand re-dissolved in a second solvent

The polymer need not be pretreated or wetted prior to contacting thesolution with the polymer. In one embodiment, the polymer is treatedwith a water-miscible organic solvent, followed by water or aqueousbuffer, prior to contacting the solution with the polymer. In anotherembodiment, the solution is contacted with dry polymer, that is, thepolymer is not wetted prior to treatment of the solution.

The solution contacted with the polymer can comprise a polar solvent andis preferably predominately, i.e. greater than 50% by volume, an acidic,basic or neutral aqueous solution or aqueous buffer. The solution canalso comprise a water-miscible polar organic solvent such as methanol,ethanol, acetonitrile, N,N-dimethylformamide, or dimethylsulfoxide, or amixture of such a solvent and water.

The solution comprising the solute of interest can further comprise oneor more additional solutes. In one embodiment, the additional solute orsolutes are more polar than the solute of interest, and, thus, adsorbmore weakly to the polymer than the solute of interest. Such anadditional solute can be desorbed from the polymer by washing thepolymer with a solvent which does not desorb the compound of interest,thereby forming a solution of the additional solute or solutes which issubstantially free of the solute of interest. A suitable solvent for thedesorption of the additional solute will typically be sufficiently polarthat it does not desorb the compound of interest.

After desorption of the additional solute or solutes, the compound ofinterest can be desorbed by washing the polymer with a suitable, i.e.,less polar, solvent. This forms a solution of the organic solute whichis substantially free from more polar solutes and is suitable for thequantitative analysis of the organic solute.

In one embodiment, the solute of interest adsorbs onto the polymer, butone or more additional solutes do not. Such an additional solute can be,for example, of sufficiently high polarity that it does not adsorb ontothe polymer. The additional solute can also comprise large molecules,for example, macromolecules such as proteins, which are unable to passthrough the pores within the polymer, and, thus, have access to only asmall fraction of the overall polymer surface area. Such molecules aretypically retained poorly, if at all, by the polymer.

In a further embodiment, the additional solute or solutes are less polarthan the solute of interest and, thus, adsorb to the polymer morestrongly than the compound of interest. The compound of interest can beweakly to moderately adsorbed or not adsorbed. If adsorbed, the soluteof interest is desorbed from the polymer by washing the polymer with asolvent of sufficient polarity that it does not desorb the additionalsolute or solutes. Thus, the compound of interest can be desorbed fromthe polymer without desorbing the other solutes.

In one embodiment, the additional solute or solutes are also analytes ofinterest. Thus a series of solutes initially present in a solution canbe separated, and solutions of each suitable for quantitative analysiscan be formed using the method of the present invention. In this case,the solution is contacted with the polymer so that the solutes adsorb tothe polymer. The solutes are then desorbed from the polymer in order ofdecreasing polarity (i.e., most polar solute first, followed by solutesof successively decreasing polarity) by washing the polymer with asequence of solvents of decreasing polarity.

Polymers, solutions and solutes which arc suitable for this methodinclude those described above. Solvents which are suitable for desorbingthe solute from the polymer will typically be polar water-miscibleorganic solvents, such as alcohols, for example, methanol, ethanol, andisopropanol, acetonitrile, acetone, and tetrahydrofuran, or mixtures ofwater and these solvents. The desorbing solvent can also be a nonpolaror moderately polar water-immiscible solvent such as dichloromethane,diethylether, chloroform, or ethylacetate. Mixtures of these solventsare also suitable. Preferred solvents or solvent mixtures must bedetermined for each individual case. A suitable solvent can bedetermined by one of ordinary skill in the art without undueexperimentation, as is routinely done in chromatographic methodsdevelopment (McDonald and Bouvier, supra, (1995); Snyder and Kirkland,Introduction to Modern Liquid Chromatography, New York: J. Wiley andSons (1974)).

The methods of the present invention can be used to prepare solutions ofa solute which are suitable for quantitative analysis via a variety oftechniques, including high performance liquid chromatography, gaschromatography, gas chromatography/mass spectrometry, and immunoassay.

The sorbent polymers used in the methods of the present invention can beprepared via standard synthetic methods. For example, apoly(divinylbenzene-co-N-vinylpyrrolidone) copolymer can be synthesizedby copolymerization of divinylbenzene and N-vinylpyrrolidone usingstandard methods of free radical polymerization which are well known inthe art. One method for forming copolymers of this type is disclosed inU.S. Pat. No. 4,382,124, issued to Meitzner et al., the contents ofwhich are incorporated herein by reference. The composition of theresulting copolymer depends upon the starting stoichiometry of the twomonomers and can be readily varied. The composition of the productcopolymer in some cases will not be substantially the same as theproportion of the starting materials, due to differences in reactivityratios among the monomers.

The invention will now be further and specifically described by thefollowing example.

Exemplification

Materials

The model solutes procainamide, acetaminophen, m-toluidine, m-toluamide,propranolol, caffeine, and 2,7-dihydroxynaphthalene were obtained fromAldrich Chemical Company (Milwaukee, Wis.), while doxepin, ranitidine,and betamethasone-17-valerate were purchased from Sigma Chemical Company(St. Louis, Mo.). The tC₁₈ bonded silica solid phase extractioncartridge was obtained from Waters Corporation (Milford, Mass.,catalogue no. WAT054960). A poly(divinylbenzene-co-N-vinylpyrrolidone)copolymer comprising about 9 mole percent N-vinylpyrrolidone wasobtained from Waters Corporation (Porapak®R). Poly(divinylbenzene) wasalso obtained from Waters (Styragel®).

Preparation of poly(divinylbenzene-co-N-vinylpyrrolidone) copolymers

To a 3000 mL flask was added a solution of 5.0 ghydroxypropylmethylcellulose (Methocel E15, Dow Chemical Co., Midland,Mich.) in 1000 mL water. To this was added a solution of 175 gdivinylbenzene (DVB HP-80, Dow), 102 g N-vinyl-2-pyrrolidone(International Specialty Products), and 1.85 g azobisisobutyronitrile(Vazo 64, Dupont Chemical Co, Wilmington, Del.) in 242 g toluene. Theresulting biphasic mixture was stirred for 30 minutes at roomtemperature using sufficient agitation to form oil droplets of thedesired micron size. The resulting suspension was then heated undermoderate agitation to 70° C. and maintained at this temperature for 20hours. The suspension was then cooled to room temperature, filtered andwashed with methanol. The filter cake was then dried in vacuo for 16hours at 80° C. The composition of the product polymer was determined byelemental analysis. Elemental analysis: N: 2.24%; mole percentN-vinylpyrrolidone: 20%.

A series of poly(divinylbenzene-co-N-vinylpyrrolidone) copolymerscomprising about 13, 14, 16, and 22 mole percent N-vinylpyrrolidone wasalso prepared by this method by varying the starting ratio of thedivinylbenzene and N-vinylpyrrolidone monomers.

A 50 mg amount of each polymer was packed into a 1 cc Sep-Pak Vac®cartridge container (Waters Corporation) having a polyethylene frit atboth the inlet and the outlet of the polymer bed to form a solid phaseextraction cartridge.

Method

Each model compound was dissolved in 20 mM phosphate buffer, pH 7, toform a solution having a concentration of 10 μg/mL.

Solid phase extraction of model solutes

The solutions of the model solutes were subjected to solid phaseextraction on solid phase extraction cartridges conditioned under twosets of conditions. In both cases the cartridge was attached to a vacuummanifold and treated with 1 mL methanol. The vacuum was set to about 4"Hg, to give a methanol flow rate of 1 mL /minute. Under the first set ofconditions ("wet conditions"), the vacuum was released when the methanollevel reached the top of the sorbent. Under the second set of conditions("dry conditions") the sorbent was allowed to dry out under vacuumfollowing conditioning with methanol. As in the first method, thecartridge was treated with 1 mL methanol, at a flow rate, under reducedpressure (4" Hg), of about 1 mL/minute. When the methanol level reachedthe top of the sorbent, the vacuum was set to 10" Hg and maintained for10 minutes to dry the polymer bed.

In both wet and dry cases, 1 mL of the model compound solution wasapplied to the cartridge at a flow rate of 1 mL/minute. A 1 mL portionof 20 mM phosphate buffer, pH 7 was then added at a flow rate of 1mL/minute. A 1 mL portion of methanol was then added at a flow rate of 1mL/minute to desorb and eluate the model compound. To the eluate wasadded an internal standard, and the model compound within the eluent wasquantitated by high performance liquid chromatography.

Results

The results are summarized in the table below, which lists polarcompounds (procainamide, acetaminophen and ranitidine), moderately polarcompounds (caffeine, m-toluamide, m-toluidine, 2,7-dihydroxynaphthalene,and propranolol) and nonpolar compounds (dipropylphthalate, doxepin andbetamethasone-13-valerate). When the sorbent was poly(divinylbenzene)all compounds except doxepin showed greater than 89% recovery when thesorbent was conditioned under wet conditions. Recovery of doxcpin, asshown, was significantly lower because this compound required greaterthan 1 mL methanol for quantitative elution. When the sorbent wastreated under dry conditions, only dipropylphthalate and betamethasonevalerate were recovered in greater than 80% yield, and recovery ofprocainamide, acetaminophen and ranitidine fell below 10%.

When the sorbent was tC₁₈ -bonded silica, each compound tested wasrecovered in high yield (>85%) under wet conditions. Allowing thesorbent to dry out had negligible effect on the recovery ofdipropylphthalate, doxepin and betamethasone valerate, but reduced theyield of caffeine, m-toluidine, m-toluamide, 2,7-dihydroxynaphthaleneand propranolol to about 13% or less.

When the sorbent was a poly(divinylbenzcne-co-N-vinylpyrrolidone)copolymer, recovery of each compound was in the range of about 80-100%when the sorbent was kept wetted. When the copolymer composition was 9mole percent N-vinylpyrrolidone, high recovery of the nonpolar compoundswas noted under both wet and dry conditions. The more polar compoundswere recovered in high yield under wet conditions but in sharply reducedyield under dry conditions. Recovery of these compounds under dryconditions dramatically increased when the N-vinylpyrrolidone componentof the copolymer was increased to about 13 mole percent or greater. Therecovery of these compounds under wet conditions was essentiallyinvariant as the copolymer composition was changed.

                                      TABLE                                       __________________________________________________________________________    Comparison of SPE recoveries for various model compounds                      Analyses performed in triplicate. DVB = divinylbenzene, NVP                   = N-vinylpyrrolidone-                                                         % NVP given as mole percent NVP.                                              __________________________________________________________________________                Percent Recovery (Average)                                                          Poly(DVB-co-NVP)                                                                       Poly(DVB-co-NVP)                                                                       Poly(DVB-co-NVP)                                                                       Poly(DVB-co-NVP)                             Poly(DVB)                                                                           9% NVP   13% NVP  14% NVP  16% NVP                          Compound    wet                                                                              dry                                                                              wet dry  wet dry  wet dry  wet dry                          __________________________________________________________________________    Procainamide                                                                              95.4                                                                             2.5*                                                                             90.8                                                                              4.9*     84.8*                                                                              94.2                                                                              84*  99.9                                                                              98.8                         Acetaminophen                                                                             98.0                                                                             2.0*                                                                             93.8*                                                                             10.4*    92.3 97.9                                                                              89.4*                                                                              104.5                                                                             104.4                        Ranitidine  95.0                                                                             5.4*                                                                             89.6                                                                              13.4*    99.7 93.5                                                                              88.4 98.3                                                                              97.6                         Caffeine    95.6*                                                                            30.8*                                                                            101.0                                                                             25.6*                                                                              96.0                                                                              95.7 98.7                                                                              96.8                                  Toluamide   98.5                                                                             54.8*                                                                            101.1                                                                             74.4*                                                                              96.5                                                                              96.3 100.8                                                                             96.4                                  Toluidine   95.6                                                                             80.7*                                                                            102.8                                                                             96.8 97.6                                                                              98.0 96.5                                                                              93.4                                  2,7-Dihydroxynaphthalene                                                                  99.5                                                                             46.5*                                                                            103.6                                                                             89.6*                                                                              96.8                                                                              96.3 95.9                                                                              94.2                                  Propranolol 92.3                                                                             55.8*                                                                            102.1                                                                             94.5 94.2                                                                              92.4                                           Dipropylphthalate                                                                         91.5                                                                             99.2                                                                             102.1                                                                             101.6                                                                              93.1                                                                              100.0                                          Doxepin     47.5+                                                                            55.4+                                                                            85.3                                                                              86.8 77.5+                                                                             78.0                                           Betamethasone-13-valerate                                                                 89.1                                                                             83.6                                                                             93.3                                                                              97.1 85.9+                                                                             89.8                                           __________________________________________________________________________                 Percent Recovery (Average)                                                    Poly(DVB-co-NVP)                                                                         Poly(DVB-co-NVP)                                                                         Poly(DVB-co-NVP)                                        20% NVP    20% NVP    22% N      tC.sub.18                       Compound     wet  dry   wet  dry   wet  dry   wet dry                         __________________________________________________________________________    Procainamide 89.1 88.3       92.3  85.7 93.3                                  Acetaminophen                                                                              96.2 94.8       93.4  96.7 102.2                                 Ranitidine   86.0 85.2       92.1  76.9 86.6                                  Caffeine     99.7 97.3  97.7 98.2  99.2 95.4  103.9                                                                             1.5*                        Toluamide    100.0                                                                              97.0  97.0 97.3  100.5                                                                              96.3  103.7                                                                             2.8*                        Toluidine    94.0 93.2  95.1 96.7  91.9 90.3  101.7                                                                             13.4*                       2,7-Dihydroxynaphthalene                                                                   96.7 95.4  95.7 94.5  94.4 90.0  102.8                                                                             0*                          Propranolol  94.1 95.4  88.5 85.5  88.0 88.3  98.2                                                                              9.3*                        Dipropylphthalate                                                                          89.5 89.3  88.9 86.0  89.1 98.5  92.2                                                                              98.4                        Doxepin      84.1 81.8  84.7 77.5  78.6 79.9  95.1                                                                              104.0                       Betamethasone-13-valerate                                                                  92.7 87.0  84.0 85.0  85.9 85.9  86.6                                                                              88.3                        __________________________________________________________________________     *Breakthrough in load/wash                                                    +Requires greater than 1 mL methanol for complete elution                

Equivalents

Those skilled in the art will recognize or be able to ascertain using nomore than routine experimentation many equivalents to the specificembodiments of the invention described herein. Such equivalents areintended to be encompassed in the scope of the following claims.

What is claimed is:
 1. A method for producing a solution comprising afirst solvent and a solute, wherein the concentration of said solute insaid solution is greater than the concentration of the solute in astarting solution comprising a second solvent and the solute, the methodcomprising the steps of:(a) contacting a volume of the starting solutionwith a water-wettable polymer formed by copolymerizing at least onehydrophobic monomer and at least one hydrophilic monomer, said polymercomprising from about 12 mole percent to about 30 mole percent of thehydrophilic monomer, whereby the solute is adsorbed onto the polymer;and (b) washing the polymer with a volume of the first solvent, saidvolume of the first solvent being less than the volume of the startingsolution in step (a), thereby desorbing the solute from the polymer andforming a solution of the solute in the first solvent wherein theconcentration of the solute in said solution is greater than theconcentration of the solute in the starting solution.
 2. The method ofclaim 1 wherein the starting solution comprises water, an aqueoussolution or a mixture of water and one or more polar organic solvents.3. The method of claim 1 wherein the first solvent is a polar,water-miscible organic solvent or a mixture of water and a polar,water-miscible solvent.
 4. The method of claim 3 wherein the firstsolvent is selected from the group consisting of acetone,tetrahydrofuran, acetonitnile, methanol, ethanol, n-propanol andisopropanol.
 5. The method of claim 1 wherein the hydrophilic monomer isN-vinylpyrrolidone.
 6. The method of claim 1 wherein the hydrophobicmonomer is divinylbenzene.
 7. The method of claim 1 wherein the polymeris a poly(divinylbenzene-co-N-vinylpyrrolidone) copolymer.
 8. The methodof claim 7 wherein the polymer comprises from about 12 mole percent toabout 30 mole percent N-vinylpyrrolidone.